Publisher Correction: Uncoupling protein 2 reprograms the tumor microenvironment to support the anti-tumor immune cycle.

In the version of this article initially published, the bars were not aligned with the data points or horizontal axis labels in Fig. 5d, and the labels along each horizontal axis of Fig. 5j-l indicating the presence (+) or absence (-) of doxycycline (Dox) were incorrectly included with the labels below that axis. Also, the right vertical bar above Fig. 7b linking 'P = 0.0001' to the key was incorrect; the correct comparison is αPD-1 versus Dox + αPD-1. Similarly, the right vertical bar above Fig. 7e linking 'P = 0.0002' to the key was incorrect; the correct comparison is αPD-1 versus Rosig + αPD-1. The errors have been corrected in the HTML and PDF versions of the article.

Uncoupling protein 2 reprograms the tumor microenvironment to support the anti-tumor immune cycle.

Immune checkpoint blockade therapy has shifted the paradigm for cancer treatment. However, the majority of patients lack effective responses due to insufficient T cell infiltration in tumors. Here we show that expression of mitochondrial uncoupling protein 2 (UCP2) in tumor cells determines the immunostimulatory feature of the tumor microenvironment (TME) and is positively associated with prolonged survival. UCP2 reprograms the immune state of the TME by altering its cytokine milieu in an interferon regulatory factor 5-dependent manner. Consequently, UCP2 boosts the conventional type 1 dendritic cell- and CD8+ T cell-dependent anti-tumor immune cycle and normalizes the tumor vasculature. Finally we show, using either a genetic or pharmacological approach, that induction of UCP2 sensitizes melanomas to programmed cell death protein-1 blockade treatment and elicits effective anti-tumor responses. Together, this study demonstrates that targeting the UCP2 pathway is a potent strategy for alleviating the immunosuppressive TME and overcoming the primary resistance of programmed cell death protein-1 blockade.

Nicotinamide N-methyltransferase sustains a core epigenetic program that promotes metastatic colonization in breast cancer.

Metastatic colonization of distant organs accounts for over 90% of deaths related to solid cancers, yet the molecular determinants of metastasis remain poorly understood. Here, we unveil a mechanism of colonization in the aggressive basal-like subtype of breast cancer that is driven by the NAD+ metabolic enzyme nicotinamide N-methyltransferase (NNMT). We demonstrate that NNMT imprints a basal genetic program into cancer cells, enhancing their plasticity. In line, NNMT expression is associated with poor clinical outcomes in patients with breast cancer. Accordingly, ablation of NNMT dramatically suppresses metastasis formation in pre-clinical mouse models. Mechanistically, NNMT depletion results in a methyl overflow that increases histone H3K9 trimethylation (H3K9me3) and DNA methylation at the promoters of PR/SET Domain-5 (PRDM5) and extracellular matrix-related genes. PRDM5 emerged in this study as a pro-metastatic gene acting via induction of cancer-cell intrinsic transcription of collagens. Depletion of PRDM5 in tumor cells decreases COL1A1 deposition and impairs metastatic colonization of the lungs. These findings reveal a critical activity of the NNMT-PRDM5-COL1A1 axis for cancer cell plasticity and metastasis in basal-like breast cancer.

SST-editing: in silico spatial transcriptomic editing at single-cell resolution.

MOTIVATION: Generative Adversarial Nets (GAN) achieve impressive performance for text-guided editing of natural images. However, a comparable utility of GAN remains understudied for spatial transcriptomics (ST) technologies with matched gene expression and biomedical image data. RESULTS: We propose In Silico Spatial Transcriptomic editing that enables gene expression-guided editing of immunofluorescence images. Using cell-level spatial transcriptomics data extracted from normal and tumor tissue slides, we train the approach under the framework of GAN (Inversion). To simulate cellular state transitions, we then feed edited gene expression levels to trained models. Compared to normal cellular images (ground truth), we successfully model the transition from tumor to normal tissue samples, as measured with quantifiable and interpretable cellular features. AVAILABILITY AND IMPLEMENTATION: https://github.com/CTPLab/SST-editing.

Multiplex analysis of intratumoural immune infiltrate and prognosis in patients with stage II-III colorectal cancer from the SCOT and QUASAR 2 trials: a retrospective analysis.

BACKGROUND: Tumour-infiltrating CD8+ cytotoxic T cells confer favourable prognosis in colorectal cancer. The added prognostic value of other infiltrating immune cells is unclear and so we sought to investigate their prognostic value in two large clinical trial cohorts. METHODS: We used multiplex immunofluorescent staining of tissue microarrays to assess the densities of CD8+, CD20+, FoxP3+, and CD68+ cells in the intraepithelial and intrastromal compartments from tumour samples of patients with stage II-III colorectal cancer from the SCOT trial (ISRCTN59757862), which examined 3 months versus 6 months of adjuvant oxaliplatin-based chemotherapy, and from the QUASAR 2 trial (ISRCTN45133151), which compared adjuvant capecitabine with or without bevacizumab. Both trials included patients aged 18 years or older with an Eastern Cooperative Oncology Group performance status of 0-1. Immune marker predictors were analysed by multiple regression, and the prognostic and predictive values of markers for colorectal cancer recurrence-free interval by Cox regression were assessed using the SCOT cohort for discovery and QUASAR 2 cohort for validation. FINDINGS: After exclusion of cases without tissue microarrays and with technical failures, and following quality control, we included 2340 cases from the SCOT trial and 1069 from the QUASAR 2 trial in our analysis. Univariable analysis of associations with recurrence-free interval in cases from the SCOT trial showed a strong prognostic value of intraepithelial CD8 (CD8IE) as a continuous variable (hazard ratio [HR] for 75th vs 25th percentile [75vs25] 0·73 [95% CI 0·68-0·79], p=2·5 × 10-16), and of intrastromal FoxP3 (FoxP3IS; 0·71 [0·64-0·78], p=1·5 × 10-13) but not as strongly in the epithelium (FoxP3IE; 0·89 [0·84-0·96], p=1·5 × 10-4). Associations of other markers with recurrence-free interval were moderate. CD8IE and FoxP3IS retained independent prognostic value in bivariable and multivariable analysis, and, compared with either marker alone, a composite marker including both markers (CD8IE-FoxP3IS) was superior when assessed as a continuous variable (adjusted [a]HR75 vs 25 0·70 [95% CI 0·63-0·78], p=5·1 × 10-11) and when categorised into low, intermediate, and high density groups using previously published cutpoints (aHR for intermediate vs high 1·68 [95% CI 1·29-2·20], p=1·3 × 10-4; low vs high 2·58 [1·91-3·49], p=7·9 × 10-10), with performance similar to the gold-standard Immunoscore. The prognostic value of CD8IE-FoxP3IS was confirmed in cases from the QUASAR 2 trial, both as a continuous variable (aHR75 vs 25 0·84 [95% CI 0·73-0·96], p=0·012) and as a categorical variable for low versus high density (aHR 1·80 [95% CI 1·17-2·75], p=0·0071) but not for intermediate versus high (1·30 [0·89-1·88], p=0·17). INTERPRETATION: Combined evaluation of CD8IE and FoxP3IS could help to refine risk stratification in colorectal cancer. Investigation of FoxP3IS cells as an immunotherapy target in colorectal cancer might be merited. FUNDING: Medical Research Council, National Institute for Health Research, Cancer Research UK, Swedish Cancer Society, Roche, and Promedica Foundation.

Prognostic impact and causality of age on oncological outcomes in women with endometrial cancer: a multimethod analysis of the randomised PORTEC-1, PORTEC-2, and PORTEC-3 trials.

BACKGROUND: Numerous studies have shown that older women with endometrial cancer have a higher risk of recurrence and cancer-related death. However, it remains unclear whether older age is a causal prognostic factor, or whether other risk factors become increasingly common with age. We aimed to address this question with a unique multimethod study design using state-of-the-art statistical and causal inference techniques on datasets of three large, randomised trials. METHODS: In this multimethod analysis, data from 1801 women participating in the randomised PORTEC-1, PORTEC-2, and PORTEC-3 trials were used for statistical analyses and causal inference. The cohort included 714 patients with intermediate-risk endometrial cancer, 427 patients with high-intermediate risk endometrial cancer, and 660 patients with high-risk endometrial cancer. Associations of age with clinicopathological and molecular features were analysed using non-parametric tests. Multivariable competing risk analyses were performed to determine the independent prognostic value of age. To analyse age as a causal prognostic variable, a deep learning causal inference model called AutoCI was used. FINDINGS: Median follow-up as estimated using the reversed Kaplan-Meier method was 12·3 years (95% CI 11·9-12·6) for PORTEC-1, 10·5 years (10·2-10·7) for PORTEC-2, and 6·1 years (5·9-6·3) for PORTEC-3. Both overall recurrence and endometrial cancer-specific death significantly increased with age. Moreover, older women had a higher frequency of deep myometrial invasion, serous tumour histology, and p53-abnormal tumours. Age was an independent risk factor for both overall recurrence (hazard ratio [HR] 1·02 per year, 95% CI 1·01-1·04; p=0·0012) and endometrial cancer-specific death (HR 1·03 per year, 1·01-1·05; p=0·0012) and was identified as a significant causal variable. INTERPRETATION: This study showed that advanced age was associated with more aggressive tumour features in women with endometrial cancer, and was independently and causally related to worse oncological outcomes. Therefore, our findings suggest that older women with endometrial cancer should not be excluded from diagnostic assessments, molecular testing, and adjuvant therapy based on their age alone. FUNDING: None.

B cells critical for outcome in high grade serous ovarian carcinoma.

Recent work has shown evidence for the prognostic significance of tumor infiltrating B cells (B-TIL) in high grade serous ovarian carcinoma (HGSOC), the predominant histological subtype of ovarian cancer. However, it remains unknown how the favorable prognosis associated with B-TIL relates to the current standard treatments of primary debulking surgery (PDS) followed by chemotherapy or (neo-)adjuvant chemotherapy (NACT) combined with interval debulking surgery. To address this, we analyzed the prognostic impact of B-TIL in relationship to primary treatment and tumor infiltrating T cell status in a highly homogenous cohort of HGSOC patients. This analysis involved a combined approach utilizing histological data and high-dimensional flow cytometry analysis. Our findings indicate that while HGSOC tumors pre-treated with NACT are infiltrated with tumor-reactive CD8+ and CD4+ TIL subsets, only B-TIL and IgA plasma blasts confer prognostic benefit in terms of overall survival. Importantly, the prognostic value of B-TIL and IgA plasma blasts was not restricted to patients treated with NACT, but was also evident in patients treated with PDS. Together, our data point to a critical prognostic role for B-TIL in HGSOC patients independent of T cell status, suggesting that alternative treatment approaches focused on the activation of B cells should be explored for HGSOC.

Immune Phenotype-Genotype Associations in Primary Clear Cell Renal Cell Carcinoma and Matched Metastatic Tissue.

Adjuvant immunotherapy has been recently recommended for patients with metastatic clear cell renal cell carcinoma (ccRCC), but there are no tissue biomarkers to predict treatment response in ccRCC. Potential predictive biomarkers are mainly assessed in primary tumor tissue, whereas metastases (METs) remain understudied. To explore potential differences between genomic alterations and immune phenotypes in primary tumors and their matched METs, we analyzed primary tumors (PTs) of 47 ccRCC patients and their matched distant METs by comprehensive targeted parallel sequencing, whole-genome copy number variation analysis, determination of microsatellite instability, and tumor mutational burden. We quantified the spatial distribution of tumor-infiltrating CD8+ T cells and coexpression of the T-cell-exhaustion marker thymocyte selection-associated high mobility group box (TOX) by digital immunoprofiling and quantified tertiary lymphoid structures. Most METs were pathologically "cold." Inflamed, pathologically "hot" PTs were associated with decreased disease-free survival, worst for patients with high levels of CD8+TOX+ T cells. Interestingly, inflamed METs showed a relative increase in exhausted CD8+TOX+ T cells and increased accumulative size of tertiary lymphoid structures compared with PTs. Integrative analysis of molecular and immune phenotypes revealed BAP1 and CDKN2A/B deficiency to be associated with an inflamed immune phenotype. Our results highlight the distinct spatial distribution and differentiation of CD8+ T cells at metastatic sites, and the association of an inflamed microenvironment with specific genomic alterations.

Poor Diagnostic Reproducibility in the Identification of Nonconventional Dysplasia in Colitis Impacts the Application of Histologic Stratification Tools.

Due to their increased cancer risk, patients with longstanding inflammatory bowel disease are offered endoscopic surveillance with concomitant histopathologic assessments, aimed at identifying dysplasia as a precursor lesion of colitis-associated colorectal cancer. However, this strategy is beset with difficulties and limitations. Recently, a novel classification criterion for colitis-associated low-grade dysplasia has been proposed, and an association between nonconventional dysplasia and progression was reported, suggesting the possibility of histology-based stratification of patients with colitis-associated lesions. Here, a cohort of colitis-associated lesions was assessed by a panel of 6 experienced pathologists to test the applicability of the published classification criteria and try and validate the association between nonconventional dysplasia and progression. While confirming the presence of different morphologic patterns of colitis-associated dysplasia, the study demonstrated difficulties concerning diagnostic reproducibility between pathologists and was unable to validate the association of nonconventional dysplasia with cancer progression. Our study highlights the overall difficulty of using histologic assessment of precursor lesions for cancer risk prediction in inflammatory bowel disease patients and suggests the need for a different diagnostic strategy that can objectively identify high-risk phenotypes.

Bone marrow haematopoiesis in patients with COVID-19.

AIMS: Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infection broadly affects organ homeostasis, including the haematopoietic system. Autopsy studies are a crucial tool for investigation of organ-specific pathologies. Here we perform an in-depth analysis of the impact of severe coronavirus disease 2019 (COVID-19) on bone marrow haematopoiesis in correlation with clinical and laboratory parameters. METHODS AND RESULTS: Twenty-eight autopsy cases and five controls from two academic centres were included in the study. We performed a comprehensive analysis of bone marrow pathology and microenvironment features with clinical and laboratory parameters and assessed SARS-CoV-2 infection of the bone marrow by quantitative polymerase chain reaction (qPCR) analysis. In COVID-19 patients, bone marrow specimens showed a left-shifted myelopoiesis (19 of 28, 64%), increased myeloid-erythroid ratio (eight of 28, 28%), increased megakaryopoiesis (six of 28, 21%) and lymphocytosis (four of 28, 14%). Strikingly, a high proportion of COVID-19 specimens showed erythrophagocytosis (15 of 28, 54%) and the presence of siderophages (11 of 15, 73%) compared to control cases (none of five, 0%). Clinically, erythrophagocytosis correlated with lower haemoglobin levels and was more frequently observed in patients from the second wave. Analysis of the immune environment showed a strong increase in CD68+ macrophages (16 of 28, 57%) and a borderline lymphocytosis (five of 28, 18%). The stromal microenvironment showed oedema (two of 28, 7%) and severe capillary congestion (one of 28, 4%) in isolated cases. No stromal fibrosis or microvascular thrombosis was found. While all cases had confirmed positive testing of SARS-CoV-2 in the respiratory system, SARS-CoV-2 was not detected in the bone marrow by high-sensitivity PCR, suggesting that SARS-CoV-2 does not commonly replicate in the haematopoietic microenvironment. CONCLUSIONS: SARS-CoV-2 infection indirectly impacts the haematological compartment and the bone marrow immune environment. Erythrophagocytosis is frequent and associated with lower haemoglobin levels in patients with severe COVID-19.